Infection of Cattle with Parainfluenza 3 Virus with Special Reference to Udder Infection

Thirty-seven strains of parainfluenza 3 virus were isolated on bovine kidney cell cultures from Japanese cattle in herds suffering from acute respiratory illness. This finding, together with the clinical and epidemiologic observations and the results of a serologic survey, indicates that in Japan, as in the United States and other countries, this virus is one of the important causes of acute respiratory disease in cattle. A significant finding in the present study is that virus was recovered from milk as well as from nasal secretions. This finding suggests an important role for milk, along with nasal discharges, in disseminating the virus among cattle. In addition, the recovery of virus from milk presents a new problem concerning this infection in cattle, in particular, the potential role of this virus in the pathogenesis of mastitis.     

Bovine Adenovirus

Two adenovirus strains were isolated in calf testicle cell cultures from blood specimens of cattle in Japan. This is the first isolation of bovine adenovirus reported in Japan. The isolates were antigenically similar to each other and distinct from the hitherto described serotypes 1, 2 and 3 of bovine adenovirus. Unfortunately, bovine adenovirus types 4 and 5 were not available for comparison, and hence, until the matter is settled, the virus will be called “Bovine adenovirus type Nagano”. Nagano virus was identified as adenovirus on the bases of the inhibitory effect of 5-iodo-2′-deoxyuridine on virus replication, ether-resistance, effect of temperature and pH on infectivity, and fine structure of the virus particle. The virus grew and formed intranuclear inclusion bodies, a characteristic of adenovirus, in bovine testicle cells but not in bovine kidney cells. The virus agglutinated rat erythrocytes very poorly, but not sheep, goat, cattle, horse, guinea pig, hamster, chicken, and mouse cells. The virus produced adenovirus group-specific antigen in cell cultures. Sero-negative calves were readily infected with the virus by the intravenous, subcutaneous, oral or intranasal routes of inoculation. The infected animals produced antibodies and showed a mild clinical reaction comprised of rhinorrhea, diarrhea and a degree of pyrexia; low-titered viremia of short duration and leukopenia were also observed. A serologic survey indicated wide-spread dissemination of the virus among Japanese cattle, but further studies are needed to determine the etiologic significance of the virus in the natural disease in cattle.     

Bovine Adenovirus

Nine strains of an adenovirus serotype were recovered in bovine testicle cell cultures from Japanese cattle suffering with an acute febrile illness accompanied by rhinorrhea and diarrhea. The isolated virus was shown to have the physicochemical properties of the adenovirus group such as the nucleic acid type, the size and ultrastructure of the virion, and the resistance to ether and chloroform. The isolated virus produced eosinophilic intranuclear inclusion bodies characteristic of adenoviruses and the group specific antigen of adenovirus in bovine testicle cell culture. According to the results of cross-neutralization tests the isolated virus represents a serological type distinct from bovine adenovirus types 1, 2 and 3 and from the Nagano virus. The isolated virus agglutinated erythrocytes of cattle, sheep, goat, guinea pig, rat, hamster and mouse, but not those of vervet monkey, horse, goose and chicken. HI test using cattle erythrocytes corroborated the results of serological typing by neutralization tests.     

Bovine Respiratory Syncytial Virus

A large epizootic of an acute respiratory disease of cattle occurred in Japan during the months from October 1968 to May 1969. A virus was recovered in primary cultures of calf kidney and testicle cells from nasal swabs of affected cattle. Neutralization tests revealed the virus to be closely related to the Long strain of human respiratory syncytial virus. The virus induced cytopathic changes including the formation of syncytia and acidophilic-cytoplasmic inclusions in calf kidney and testicle cell cultures. A calf inoculated with the virus by the respiratory route developed an illness resembling the natural disease. Most cattle clinically diagnosed as having the disease showed significant rises of neutralizing antibody titer for the isolated virus, whereas none or only small fractions of those animals showed serological evidence for recent infection with bovine ephemeral fever virus, infectious bovine rhinotracheitis virus, Ibaraki virus, bovine diarrhea virus, bovine adenovirus Type 7 and parainfluenza virus Type 3. Neutralization tests on paired sera revealed a wide dissemination of the isolated virus among cattle in many areas of the country during the epizootic. All these findings leave no doubt that the epizootic was caused by bovine respiratory syncytial virus. This is the first study that ever shows the presence of infection of cattle with this virus in Japan.     

Molecular cloning and expression analysis of bovine programmed death‐1

Recent work has shown that PD‐1, an immune inhibitory receptor, is involved in mechanisms for down‐regulating immune responses during tumor progression or chronic viral infection. However, in the case of bovine diseases, there have been no reports on this molecule due to lack of information about bovine PD‐1. In this study, we performed identification and preliminary characterization of the bovine PD‐1 gene in two breeds of cattle. We cloned full cDNA sequences encoding for PD‐1 from both Holstein‐Friesian and Japanese Black breeds, and found that both of the genes encoded a 282‐amino acid protein, which had a signal sequence, transmembrane domain and an immunoreceptor tyrosine‐based inhibitory motif. This bovine PD‐1 showed 72.9% and 65.6% homology to human and mouse PD‐1, respectively, both of which have been well characterized and documented. Quantitative real‐time PCR analysis showed that bovine PD‐1 is expressed predominantly in T‐cells (such as CD4⁺ and CD8⁺ cells) and among PBMCs, and is strongly upregulated on T‐cell stimulation via ConA. A limited number of cattle were tested yet, as expected, the degree of PD‐1 mRNA expression in CD4⁺ and CD8⁺ T‐cells was greater in cattle with bovine leukemia virus‐induced lymphoma than in uninfected cattle. Further studies to characterize the functions of bovine PD‐1 are therefore warranted, in order to elucidate the mechanism of the immunosuppression associated with progression of several diseases and therapy in cattle.     

Preliminary studies of the complement fixation test to confirm the diagnosis of bovine ephemeral fever

A strain of bovine ephemeral fever (BEF) virus isolated in China in 1976 was adapted to growth in tissue cultures. A baby hamster kidney complement fixing (CF) antigen, stable at -20 degrees C for at least 120 days, was prepared from the BEF virus grown in tissue culture and used to test bovine sera for antibodies to that virus. CF antibodies were detected in all of 31 cattle after convalescence from experimental infection with BEF virus, in 208 (98%) of 213 cattle observed to have shown clinical ephemeral fever in an epidemic, in 96 cattle in these herds which did not show clinical signs of ephemeral fever and 16 cattle from herds in northern China outside the epidemic area. The CF antibodies to BEF virus were found to persist in 34 (89%) of 38 cattle which were bled 6 years after natural exposure to ephemeral fever. The CF antigen is economical to prepare and is suitable to differentiate ephemeral fever from other viral infections with which it could possibly be confused on clinical appearance.     

Localization of the locus responsible for Chediak-Higashi syndrome in cattle to bovine chromosome 28

Chediak-Higashi syndrome in Japanese black cattle is a hereditary disease with prolonged bleeding time and partial albinism. In the present study, we mapped the locus responsible for the disease (CHS) by linkage analysis using microsatellite genotypes of paternal half-sib pedigrees obtained from commercial herds. Analysis revealed significant linkage between the CHS locus and marker loci on the proximal end of bovine chromosome 28. The CHS locus was mapped on the region incorporating the microsatellite markers BMC6020, BM2892, and RM016 with recombination fraction 0 and lod score 4.9-11.2. We also assigned the bovine CHS1/LYST, the homologue of the gene responsible for human Chediak-Higashi syndrome, to bovine chromosome 28 using a bovine/murine somatic cell hybrid panel. These findings suggest that a mutation in the CHS1/LYST gene is likely to be responsible for Chediak-Higashi syndrome in Japanese black cattle.     

Cloning of bovine LYST gene and identification of a missense mutation associated with Chediak-Higashi syndrome of cattle

An inheritable bleeding disorder with light coat color caused by an autosomal recessive gene has been reported in a population of Japanese black cattle. The disease has been diagnosed as Chediak-Higashi Syndrome (CHS) of cattle which correspond to a human inheritable disorder caused by mutation in LYST gene. To characterize the molecular lesion causing CHS in cattle, cDNAs encoding bovine LYST were isolated from a bovine brain cDNA library. The nucleotide and deduced amino acid sequences of bovine LYST had 89.6 and 90.2% identity with those of the human LYST gene, respectively. In order to identify the mutation within the LYST gene causing CHS in cattle, cDNA fragments of the LYST gene were amplified from an affected animal by RT-PCR and their nucleotide sequences were completely determined. Notably, a nucleotide substitution of A to G transition, resulting in an amino acid substitution of histidine to arginine (H2015R) was identified in the affected animal. The presence of the substitution was completely corresponding with the occurrence of the CHS phenotype among 105 members of pedigrees of the Japanese black cattle and no cattle of other populations had this substitution. These findings strongly suggested that H2015R is the causative mutation in CHS of Japanese black cattle. Received: 25 May 1999 / Accepted: 26 July 1999     

Significance of Neospora caninum in British dairy cattle determined by estimation of seroprevalence in normally calving cattle and aborting cattle

A case control study was conducted to evaluate the significance of Neospora caninum infections in cattle in England and Wales. The prevalence of N. caninum in normally calving cattle (the control group; n = 418) and aborting cattle (n = 633) was estimated using a commercial antibody-detection ELISA. Prevalence estimates for bovine virus diarrhoea virus, infectious bovine rhinotracheitis virus and Leptospira hardjo were also obtained by serology. The prevalence of N. caninum was significantly higher (P < 0.0001) in the aborting group (18%; 95% confidence interval: 15%, 21%) than in the control group (6%; 95% confidence interval: 4%, 8%); the latter is the first estimate, to date, of the national seroprevalence of N. caninum in dairy cattle in England and Wales. Prevalence estimates for bovine virus diarrhoea virus, infectious bovine rhinotracheitis virus and L. hardjo were not found to be higher in the aborting cattle than in the control group. With N. caninum, a strong association between seropositivity and abortion was found, with seropositive cows being 3.5-times more likely to abort than seronegative cows (odds ratio = 3.49; 95% confidence interval: 2.16, 5.69). Furthermore, 12.5% of abortions in dairy cattle in England and Wales may be attributable to N. caninum, as indicated by estimation of the population aetiological fraction.     

Antibodies to bovine bacterial and viral pathogens in pronghorns in Alberta, 1983

Sera from 210 pronghorns (Antilocapra americana) ranging in southeastern Alberta were tested for antibodies to disease agents present in indigenous cattle. No antibodies to Brucella abortus, Leptospira interrogans serovars pomona, hardjo, or grippotyphosa, or infectious bovine rhinotracheitis virus were found. Antibodies at prevalences of 43.8% and 49.2% were detected to bovine virus diarrhea (BVD) and parainfluenza type 3 (PI-3) viruses, respectively. The much higher prevalence of BVD virus antibodies in cattle than in pronghorns, and the occurrence of clinical bovine PI-3 infection in the study area, suggest that cattle may be a source of infection to the pronghorns.     

Identification of newly isolated Babesia parasites from cattle in Korea by using the Bo-RBC-SCID mice

Attempts were made to isolate and identify Korean bovine Babesia parasite. Blood samples were collected from Holstein cows in Korea, and Babesia parasites were propagated in SCID mice with circulating bovine red blood cells for isolation. The isolate was then antigenically and genotypically compared with several Japanese isolates. The Korean parasite was found to be nearly identical to the Oshima strain isolated from Japanese cattle, which was recently designated as Babesia ovata oshimensis n. var. Haemaphysalis longicornis was the most probable tick species that transmitted the parasite.     

Occurrence of haploid cells bearing Y chromosomes in bovine embryos fertilized in vitro

Haploid cells bearing Y chromosomes were observed in bovine embryos derived from in vitro fertilization of in vitro-matured follicular oocytes in Japanese Black cattle. The androgenic embryos described in this article might be caused by abnormality in the formation of the female pronucleus.     

Typing of pestiviruses from eland in Zimbabwe

Pestiviruses were isolated from three eland (Taurotragus oryx) in Zimbabwe. The viruses were characterised by typing with monoclonal antibodies and by partial genetic sequencing. All were similar to bovine viral diarrhea viruses commonly isolated from cattle. This suggests that bovine viral diarrhea virus can spread from cattle to eland.     

Expressed 2-5A synthetase genes and pseudogenes in the marine sponge Geodia barretti

In cattle, bovine leukocyte antigens (BoLAs) have been extensively used as markers for bovine diseases and immunological traits. In this study, we sequenced alleles of the BoLA class II loci, BoLA-DRB3 and BoLA-DQA1, from 650 Japanese cattle from six herds [three herds (507 animals) of Japanese Black cattle and three herds (143 animals) of Holstein cattle] using polymerase chain reaction-sequence-based typing (PCR-SBT) methods. We identified 26 previously reported distinct DRB3 alleles in the two populations: 22 in Japanese Black and 17 in Holstein. The number of DRB3 alleles detected in each herd ranged from 9 to 20. Next, we identified 15 previously reported distinct DQA1 alleles: 13 in Japanese Black and 10 in Holstein. The number of alleles in each herd ranged from 6 to 10. Thus, allelic divergence is significantly greater for DRB3 than for DQA1. A population tree on the basis of the frequencies of the DRB3 and DQA1 alleles showed that, although the genetic distance differed significantly between the two cattle breeds, it was closely related within the three herds of each breed. In addition, Wu-Kabat variability analysis indicated that the DRB3 gene was more polymorphic than the DQA1 gene in both breeds and in all herds, and that the majority of the hypervariable positions within both loci corresponded to pocket-forming residues. The DRB3 and DQA1 heterozygosity for both breeds within each herd were calculated based on the Hardy-Weinberg equilibrium. Only one Japanese Black herd showed a significant difference between the expected and observed heterozygosity at both loci. This is the first report presenting a detailed study of the allelic distribution of BoLA-DRB3 and -DQA1 genes in Japanese Black and Holstein cattle from different farms in Japan. These results may help to develop improved livestock breeding strategies in the future.     

Diseases of wapiti utilizing cattle range in southwestern Alberta

Specimens from 28 wapiti (Cervus elaphus canadensis) were collected by hunters in southwestern Alberta in 1984. Various tests were performed to detect infections and conditions that could affect cattle sharing the range or cause disease in wapiti. Serum antibodies were present against leptospiral serovars autumnalis (25%), bratislava (4%), and icterohaemorrhagiae (8%), and the viruses of bovine virus diarrhea (52%), infectious bovine rhinotracheitis (45%), and parainfluenza type 3 (13%). No serological evidence of bovine respiratory syncytial virus, Brucella, Anaplasma, bluetongue virus, or epizootic hemorrhagic disease virus was found, nor were any lesions of vesicular diseases, necrotic stomatitis or nutritional myopathy evident. Focal interstitial nephritis and sarcocystosis were diagnosed histologically in 40% and 75%, respectively, of the wapiti tested. The prevalence of giant liver flukes (Fascioloides magna) was 50% and of lungworms (Dictyocaulus viviparus) 32%. Leptospiral serology on cattle in the area did not indicate that wapiti or cattle were a serious source of infection to each other. The giant liver fluke was the parasite most likely to be amplified by wapiti for cattle. Within the limits of this study, the results indicated that wapiti in the Waterton area do not pose a disease threat to the cattle with which they range, but periodic observational studies in these wapiti would be a useful means of early detection of any changes in the interspecies relationship.     

Diagnosis of bovine respiratory syncytial virus infection by complement fixation test.

The complement fixation test by the microtiter method was applied to the serological diagnosis of bovine respiratory syncytial (RS) virus infection. When used as complement fixing antigens, untreated infected cell culture fluid, fluorocarbon-treated, and ether-treated materials showed no differences in antigenicity among them. The complement fixing antigenicity of bovine RS virus appeared in bovine kidney and Vero cell cultures for the first time 4 days after inoculation. Both the infectivity and complement fixing antigenicity reached a maximum 6 days after inoculation. In detecting complement fixing antibody from infected cattle, the most outstanding specific reaction was obtained when 5% fresh normal calf serum had been added to the diluent of complement. Neutralizing and complement fixing antibodies were examined in serum samples from two cattle in the course of experimental infection. It was found that both antibodies turned to be positive 2 weeks after inoculation. There was a linear correlation between neutralizing and complement fixing antibody titers, when serum samples from 40 natural cases were tested in the acute and convalescent stages. In addition, common antigenicity was demonstrated between the virus of bovine origin and the Long strain of human RS virus by complement fixation test.     

Identification and characterization of bovine programmed death‐ligand 2

Previous reports from this group have indicated that the immunoinhibitory programmed death (PD)‐1 receptor and its ligand, PD‐L1, are involved in the mechanism of immune evasion of bovine chronic infection. However, no functional analysis of bovine PD‐L2 in cattle has been reported. Thus, in this study, the molecular function of bovine PD‐L2 was analyzed in vitro. Recombinant PD‐L2 (PD‐L2‐Ig), which comprises an extracellular domain of bovine PD‐L2 fused to the Fc portion of rabbit IgG1, was prepared based on the cloned cDNA sequence for bovine PD‐L2. Bovine PD‐L2‐Ig bound to bovine PD‐1‐expressing cells and addition of soluble bovine PD‐1‐Ig clearly inhibited the binding of PD‐L2‐Ig to membrane PD‐1 in a dose‐dependent manner. Cell proliferation and IFN‐γ production were significantly enhanced in the presence of PD‐L2‐Ig in peripheral blood mononuclear cells (PBMCs) from cattle. Moreover, PD‐L2‐Ig significantly enhanced IFN‐γ production from virus envelope peptides‐stimulated PBMCs derived from bovine leukemia virus‐infected cattle. Interestingly, PD‐L2‐Ig‐induced IFN‐γ production was further enhanced by treatment with anti‐bovine PD‐1 antibody. These data suggest potential applications of bovine PD‐L2‐Ig as a therapy for bovine diseases.     

Single Pathogen Challenge with Agents of the Bovine Respiratory Disease Complex

Bovine respiratory disease complex (BRDC) is an important cause of mortality and morbidity in cattle; costing the dairy and beef industries millions of dollars annually, despite the use of vaccines and antibiotics. BRDC is caused by one or more of several viruses (bovine respiratory syncytial virus, bovine herpes type 1 also known as infectious bovine rhinotracheitis, and bovine viral diarrhea virus), which predispose animals to infection with one or more bacteria. These include: Pasteurella multocida, Mannheimia haemolytica, Mycoplasma bovis, and Histophilus somni. Some cattle appear to be more resistant to BRDC than others. We hypothesize that appropriate immune responses to these pathogens are subject to genetic control. To determine which genes are involved in the immune response to each of these pathogens it was first necessary to experimentally induce infection separately with each pathogen to document clinical and pathological responses in animals from which tissues were harvested for subsequent RNA sequencing. Herein these infections and animal responses are described.     

一株牛源阿卡斑病毒的分离鉴定

阿卡斑病毒(Akabane virus,AKV)是能引起牛、绵羊、山羊流产、早产、新生胎儿畸形的虫媒性RNA病毒。为了解家畜虫媒病毒在我国西南边境地区的分布和流行情况,本研究对中缅边境西盟县的52份牛抗凝血和140份血清(牛70份、羊70份)中的蓝舌病病毒(Bluetongue virus,BTV)、鹿流行性出血热病毒(Epizootic hemorrhagic disease virus,EHDV)、AKV等虫媒病毒进行检测与分离,通过ELISA及qRT-PCR方法检测病毒,通过核酸阳性抗凝血接种BHK细胞传代以分离病毒,通过设计特异性引物,扩增分离毒株S基因721bp片段及M基因816bp片段,通过克隆测序及中和试验以鉴定病毒,最终从38号牛的抗凝血中分离到一株AKV,TCID50为10-3.5/0.1mL,经比对,分离株S片段与日本KS-2/Mo/06毒株亲缘关系最近,核苷酸同源性为97.66%,M片段与中国DHL10M110毒株亲缘关系最近,核苷酸同源性为96.56%。本研究首次报告了从云南边境地区牛群中分离到AKV,证实了西南边境存在AKV的流行,为AKV在我国的流行病学和边境地区疫病风险防控提供了重要参考及有力依据。